Project Summary/Abstract Late-onset Alzheimer?s disease (AD) is a neurodegenerative disorder characterized by progressive loss of memory and other cognitive functions. While the pathology and clinical course of AD is well-documented, there are presently no effective disease-modifying therapies or cures in existence. With the incidence of AD expected to climb over the next half century, it is important to further investigate the molecular and cellular mechanisms underlying AD. Genome-wide association studies have identified genetic variants that are highly associated with AD, but their contributions to AD have not been completely elucidated. Bridging Integrator 1 (BIN1) is one of these genes; although it is known to play a role in cellular processes such as endocytosis and membrane trafficking, little is known about how the gene is regulated and how it impacts AD pathogenesis. Further, BIN1 is known to be expressed in neurons, oligodendrocytes, and microglia, yet few studies have focused on microglia-specific BIN1 expression in AD. Our lab has identified a DNA regulatory region, called an enhancer, in microglia upstream of the BIN1 promoter that also contains an important AD risk variant. The central hypothesis of this proposal is that the expression of BIN1 is regulated by the putative BIN1-associated enhancer and that an AD variant within the enhancer modulates gene expression. This proposal describes an in vitro system that will systematically screen the putative microglial BIN1 enhancer for functional elements using CRISPR-mediated genome editing strategies. These studies will provide an idea of which non-coding regions of the genome are important for controlling BIN1 expression in microglia. Finally, the effect of an AD risk variant on gene expression will be studied using a novel in vivo enhancer-reporter assay. Understanding gene regulatory mechanisms in microglia is an important step towards fully defining the cellular and molecular mechanisms underlying AD risk. This proposal addresses a growing need to approach diseases from the perspective of genomics and gene regulation and will provide proof of concept for a general approach linking genetic variants with functional consequences in a cell- and disease- specific context. The proposed research will take place in the Glass Laboratory at UCSD. The lab has expertise in tissue-resident macrophage gene regulation and is composed of a diverse group of scientists whose specialties ranges from bioinformatics to cell biology. Through graduate coursework, mentorship, and hands-on learning, Bethany will gain experience in approaching large datasets from a quantitative perspective and will learn cutting-edge wet lab techniques; these skills will be valuable for the completion of the proposed research and for Bethany?s future career as a physician-scientist.